The success of gene therapies and the movement toward treatments for prevalent diseases is creating dramatically higher demand for viral vectors than can be currently met with existing capacity. While expansion can help meet some of this need, it is equally important that the efficiency and productivity of current processes be improved. It benefits the industry to enable more doses to be produced in intensified processes with smaller equipment. A comprehensive DoE strategy with a platform approach can be powerful. In combination with collaborations like the one between ABL and Polyplus the payout of productivity, safety and efficacy in viral vector manufacturing is as clear as the impact of working with quality raw materials across the process. As processes become further optimized, the industry will see higher generation of functional virus in a smaller footprint with less reagent and media. For upstream optimization of the transient transfection process, ABL evaluated the performance of PEIPro and FectoVIR-AAV with the Polyplus team. A DoE study was implemented using small-scale (125-mL) shake flasks to identify the CPPs. Further DoE runs were then performed in small-scale bioreactors (Ambr®250 modular system from Sartorius with up to eight vessels), and the process was then scaled up.